1CIU

THERMOSTABLE CGTASE FROM THERMOANAEROBACTERIUM THERMOSULFURIGENES EM1 AT PH 8.0.

1CIU の概要

• エントリーDOI: 10.2210/pdb1ciu/pdb
• 分子名称: CYCLODEXTRIN GLYCOSYLTRANSFERASE, CALCIUM ION (3 entities in total)
• 機能のキーワード: thermostable, glycosidase
• 由来する生物種: Thermoanaerobacterium thermosulfurigenes
• 細胞内の位置: Secreted: P26827
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 75578.23

構造登録者

Knegtel, R.M.A.,Dijkstra, B.W. (登録日: 1995-10-23, 公開日: 1996-03-08, 最終更新日: 2024-02-07)

主引用文献

Knegtel, R.M.,Wind, R.D.,Rozeboom, H.J.,Kalk, K.H.,Buitelaar, R.M.,Dijkhuizen, L.,Dijkstra, B.W.
Crystal structure at 2.3 A resolution and revised nucleotide sequence of the thermostable cyclodextrin glycosyltransferase from Thermonanaerobacterium thermosulfurigenes EM1.
J.Mol.Biol., 256:611-622, 1996

PubMed Abstract: The crystal structure of the cyclodextrin glycosyltransferase (CGTase) from the thermophilic microorganism Thermoanaerobacterium thermosulfurigenes EM1 has been elucidated at 2.3 A resolution. The final model consists of all 683 amino acid residues, two calcium ions and 343 water molecules, and has a crystallographic R-factor of 17.9% (Rfree 24.9%) with excellent stereochemistry. The overall fold of the enzyme is highly similar to that reported for mesophilic CGTases and differences are observed only at surface loop regions. Closer inspection of these loop regions and comparison with other CGTase structures reveals that especially loops 88-95, 335-339 and 534-539 possibly contribute with novel hydrogen bonds and apolar contacts to the stabilization of the enzyme. Other structural features that might confer thermostability to the T. thermosulfurigenes EM1 CGTase are the introduction of five new salt-bridges and three Gly to Ala/Pro substitutions. The abundance of Ser, Thr and Tyr residues near the active site and oligosaccharide binding sites might explain the increased thermostability of CGTase in the presence of starch, by allowing amylose chains to bind non-specifically to the protein. Additional stabilization of the A/E domain interface through apolar contacts involves residues Phe273 and Tyr187. No additional or improved calcium binding is observed in the structure, suggesting that the observed stabilization in the presence of calcium ions is caused by the reduced exchange of calcium from the protein to the solvent, rendering it less susceptible to unfolding. The 50% decrease in cyclization activity of the T. thermosulfurigenes EM1 CGTase compared with that of B. circulans strain 251 appears to be caused by the changes in the conformation and amino acid composition of the 88-95 loop. In the T. thermosulfurigenes EM1 CGTase there is no residue homologous to Tyr89, which was observed to take part in stacking interactions with bound substrate in the case of the B. circulans strain 251 CGTase. The lack of this interaction in the enzyme-substrate complex is expected to destabilize bound substrates prior to cyclization. Apparently, some catalytic functionality of CGTase has been sacrificed for the sake of structural stability by modifying loop regions near the active site.

PubMed: 8604143
DOI: 10.1006/jmbi.1996.0113

実験手法

X-RAY DIFFRACTION (2.3 Å)